CH645728A5 - PROCESS FOR DETERMINING THE CONTENT OF SUBSTANCES CONTAINING GLUCOSIDIC BONDS ASSOCIATED WITH TUMORS, APPLICATION OF THIS PROCESS AND NECESSARY FOR ITS IMPLEMENTATION. - Google Patents

Description

The present invention relates to the determination of the content of substances containing glucosidic linkages associated with tumors, i.e. substances related to sugar, including glucoproteins, glucopeptides, glucolipids and sugars (hereinafter abbreviated as TAG) in a mood sample of the organism in mammals, more particularly TAGs which increase as undifferentiated cells, in particular tumor or cancer cells, proliferate, and a method for diagnosing cancer based on this determination.

A method for determining the content of specific glucoproteins in a body humor sample has been used to date in cancer patients. This method uses the antigen character which is mainly possessed by the protein portion of glucoproteins. For example, the content of c ^ -fetoprotein and the content of carcinoembryonic antigen (hereinafter referred to as CEA) are determined for the diagnosis of primary hepatoma and cancers of the digestive organs, in particular cancer of the rectum, etc. [see “Igaku no Ayumi”, volume 106, N ° 5, 5th special edition 40 of Saturday, pp. 235-250 (1978)]. However, these methods are not satisfactory, since their application is limited to certain types of cancers and tumors.

On the other hand, no diagnostic method is currently known using the specificity of binding of the TAG sugar residue. 45 There has long been a need for an inexpensive and simple quantitative methodology to determine TAG levels in the moods of the organism, which may have wide applications.

Taking into account the fact that the moods of the body of individuals or cancer patients contain TAGs produced by undifferentiated cells (most often cancer cells) and released in the body's moods and that TAGs are extremely different from the substances containing glucosidic bonds produced by differentiated cells (usually normal cells) and released in the body's moods 55 in the chemical structure of the sugar chain, its length and the composition of the residues sugar, the licensee has carried out extensive research which has led to the discovery that TAGs have a galactose residue- (ßl - »3 or ßl -> - 4) -N-acetyIglucosamine and / or -acetylgalactosamine terminal and bind to specifi-6o only to lectins, and that the TAG content in a sample of the body's humor can be determined by reaction of the TAGs with a lectin. The TAG content indicates the presence or absence of cancer cells, their degree of proliferation and their increase or decrease, and allows to successfully establish the diagnosis of various cancers or tumors. The invention is based on the above discovery.

The subject of the invention is therefore a method for determining the TAG content in a sample of the organism's humor, which is

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characterized in that the TAGs present in a sample of the organism are reacted with a lectin to form a TAG-lectin complex and the amount of the TAG-lectin complex or of unprocessed lectin is measured.

The invention will be better understood on reading the description which will follow with reference to the figures of the appended drawings in which:

fig. 1 graphically represents the quantities of TAG determined according to Example 1, and FIG. 2 graphically represents the quantities of TAG determined according to Example 2.

Various moods of the organism can be used in the invention. Examples of suitable moods include blood, tissue moods, lymph, hydrothorax, ascites, amniotic fluid, gastric juice, urine, pancreatic juice, cerebrospinal fluid, saliva, etc. Among these, blood is preferred, in particular in the form of serum or plasma.

The amount of the organism's mood for the samples is usually about 1 to 10 ml, preferably 2 to 5 ml.

According to the invention, the determination of the TAG content in a body humor sample can be carried out either by isolating or by separating the TAGs from the body humor, by making them react with a lectin. to form a TAG-lectin complex and by measuring the amount of the TAG-lectin or unprocessed lectin complex after separation, either by adding a labeled lectin to the body's mood sample directly to form a TAG-lectin complex labeled, separating the resulting complex or the untransformed lectin and measuring its amount.

To separate the TAG fraction from a body sample, one can use conventional techniques for the extraction or separation of substances containing glucosidic bonds, such as salting out, precipitation, extraction by solvents, centrifugation. , dialysis, molecular sieves, enzyme inactivation, etc., or a combination of two or more of them. For example, the TAG fraction can be obtained by adding an amount, effective for precipitation or denaturation, of sulfosalicy-lic acid, trichloroacetic acid or zinc sulfate to serum or blood plasma, or by heating serum or blood plasma to precipitate albumin, immunoglobulin, etc., by filtering out the precipitated proteins and dialyzing the filtrate.

In cases where a labeled lectin is used, the organism's moods other than blood can be used as such as a test sample (hereinafter sample). It is preferred, however, to add a protein with a low sugar content such as beef sebuminum (BSA), etc., as a protective protein to avoid denaturation of the sample and at the same time initiate the reaction with the lectin. In particular, an appropriate amount of the protective protein is added to samples after separation of albumins, immunoglobulins, etc., since this allows determination under more controlled conditions. When using blood samples, it is possible to use as a sample the blood serum obtained by the conventional technique for collecting blood serum, or blood plasma obtained by the conventional technique for collecting blood plasma by means of an anticoagulant such as heparin, EDTA, citric acid, etc .; blood plasma collected using heparin as an anticoagulant is preferred. The samples can be diluted with water or with suitable buffered aqueous solutions, if necessary or desired, for example when the TAG content is relatively high, as in ascites, etc.

According to the invention, lectins can be used which can combine specifically with galactose- (ßl - <• 3 or ßl -> 4) -N-acetylglucosamine and / or -acetylgalactosamine as described in "JBC", 250, pp. 8518-8523 (1975); "Biochem. Biophys. Res. Comm. ”, 62, p. 144 (1975); "Z. Immunitaetsforch. ”, 138, pp. 423-433 (1969); "Br. J. Exp. Pathol. ”, 27, pp. 228-236 (1946); "Proc. Nati. Acad. Sci. USA ”, 75, N ° 5, pp. 2215-2219 (1978); "Biochemis-try", 13, pp. 196-204 (1974); "Carbohydrate Research", vol. 51,

pp. 107-118 (1976), etc., e.g. peanut lectin, castor lectin (Ricinus communis), etc.

■ The substances which can be used to label lectins and which therefore make it possible to detect them by radiometric, fluorometric, colorimetric and similar techniques are enzymes, fluorescent substances and radioactive substances.

Examples of suitable enzymes that may be mentioned include glu-coamylase, glucose oxidase, peroxidase, alkaline phosphatase, β-galactosidase, hemeoctapeptide, etc., and their active fragments. Examples of suitable fluorescent materials include fluorescein, fluorescein isothiocyanate, rhodamine, dansyl chloride (5-dimethylamino-1-naphthalenesulfonyl chloride), etc. Examples of suitable radioactive substances include radioactive iodine, tritium, etc. To label lectins with the labeling substances described above, any conventional technique used to label known proteins, such as antigens and antibodies, can be used with enzymes, fluorescent substances and radioactive substances. For example, lectins can be labeled with radioactive isotopes according to the method described by Kazuo Shizunome and Yuichi Kumahara in "Radioimmunoassay", pp. 45-46 (1978), published by Asakura Publishing Co., Tokyo.

In the process of the invention, use is preferably made of labeled or unlabeled lectins in admixture with suitable solvents. Any solvents which do not denature the labeled or unlabeled lectins can be used, for example physiological saline, 0.1 M tris-HCl buffer solution (pH about 7.5), phosphate buffer solution 0, 1 M (pH about 7.4), etc. The quantity of lectin contained in the solvent or diluent can be chosen according to the agglutination index (which is defined by the final or maximum dilution of samples diluted in series in report 2), the type of labeling substance or substances to be determined, etc., and is usually about 0.01 to 100 y / ml, preferably 0.03 to 40 y / ml. The above labeled or unlabeled lectin solutions can be further diluted.

To carry out the process of the invention, a predetermined amount of the fraction of the TAG or of the humor of the organism itself is mixed with a given amount of lectin or labeled lectin, respectively, and left rest the resulting mixture to allow it to react at approximately 45 ° C or less, preferably approximately 4 to 40 ° C, more particularly approximately 20 to 40 ° C. The unlabeled or labeled TAG-lectin complex can be separated from the reaction mixture , or unreacted or labeled unreacted lectin, by any conventional separation technique, for example chromatography, electrophoresis, salting out, fractionation, dialysis, gel filtration, adsorption, or a combination of these techniques or separation method using agar, agarose or polyacrylamide gel.

More specifically, to separate the unlabeled or unlabeled lectin from the reaction mixture, a suitable amount of a precipitating agent of the TAG-lectin complex can be added to the above reaction mixture and the resulting complex separated, for example by centrifugation , and thus obtaining the fraction of lectin labeled or unlabelled not transformed as supernatant. Typical examples of precipitants are po-lyethylene glycol, ammonium sulfate (saturated), ethacridine lactate monohydrate (acrinol), etc. The centrifugation conditions can be chosen suitably according to the precipitants used. For example, when polyethylene glycol is used as the precipitant, it is preferred to carry out centrifugation at approximately 1000 G for approximately 30 to 60 min.

In cases where the labeled or unlabelled TAG-lectin complex is isolated from unreacted lectin, the complex can be easily separated from the untransformed lectin using the difference in diffusion rates of the complex and the unprocessed lectin on agar, agarose or polyacrylamide gel. More particularly, when the reaction mixture is deposited on the gel in a container, the unprocessed lectin diffuses into the gel,

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while the TAG-lectin complex remains on the gel, that is to say outside the gel, without diffusing. Thus, they can be separated from one another easily in a conventional manner. There is no limitation in the preparation of gels and the techniques used in conventional manner can be used for this purpose. For example, a certain amount of agar, agarose or polyacrylamide is added to a diluent which does not cause protein denaturation, such as distilled water, citrate or tris-HCl buffer solution (pH approximately 7, 5), etc., then heat to 60-80 ° C with gentle agitation to dissolve, and pour the solution into a suitable container, such as a test tube, and allow to cool so that the mixture becomes jelly . The concentration of the gel is chosen according to the size (for example molecular weight, stereostructure, etc.) of the labeling substance and of the labeled TAG-lectin complex. Gels of about 0.4 to 2.0% by weight, preferably 0.7 to 1.0% by weight, are commonly used in the invention. The resulting gel may contain a preservative, if necessary or if desired. The gu gel surface thus prepared can be flat or concave, the latter form being preferred, since the complexes to be formed do not adhere to the wall of the container.

The quantity of labeled or unlabeled TAG-lectin complex, or of untransformed labeled or unlabeled lectin, is measured by the conventional technique and the content of TAG in the mood of the organism can be calculated from the value obtained. .

Various techniques can be used to measure the amount of residual unlabeled lectin. However, it is preferred to add to the samples a substance capable of reacting specifically with the lectin to agglutinate or precipitate it and observe the change visually, or alternatively by measuring the optical density using optical analysis. Examples of suitable lectin-binding substances include glucoproteins having a galactose- (ßl - »3 or ßl - * 4) -N-acetylglucosamine and / or -acetylgalactosamine residue, e.g. rabbit erythrocytes, sheep erythrocytes treated with neuraminidase, etc., Se-phadex, glass beads, etc., coated with the glucoproteins described above.

In practice, this method can be implemented in the following manner. The reaction mixture described above is diluted in series with a 2X dilution using a diluent, e.g. 0.15 M phosphate buffer solution, saline solution, etc., and aliquots of the diluted solution on V-shaped or U-shaped plates or on glass plates or in small test tubes or the like, and each sample is mixed with a substance capable of specifically agglutinating the lectin by shaking and the plates are left to stand or analogues bringing the mixture to 45 ° C or lower, preferably 4 to 40 ° C, for 30 min or more, preferably 60 to 90 min, to observe the final or maximum dilution at which agglutination still occurs. This maximum dilution is defined as the agglutination index. The lectins which can be used in the invention have substantially the same agglutination index.

In cases where labeled lectins are used, the determination of the TAG content can be carried out by an appropriate method for the measurement of substances labeled for lectins. For example, after separation of the TAG-lectin complex and the untransformed labeled lectin, the quantity of labeled TAG-lectin complex or of untransformed labeled lectin can be determined by choosing an appropriate support for an enzyme, this support can be analyzed by a colorimetric or fluorimetric method and determining the activity of the enzyme when the lectin is labeled with it; determining the intensity of fluorescence when the lectin is labeled with a fluorescent substance; or by determining radioactivity when the lectin is labeled with a radioactive substance. For example, when an enzyme, for example a peroxidase, is used to label the lectin, a predetermined amount of the reaction mixture is added dropwise to the gel surface in a test tube and allowed to stand near the room temperature, preferably 4-

25 ° C, for 1 to 48 h. A given quantity of a suitable substrate for the enzyme (for example hydrogen peroxide) is then added to the reaction mixture thus treated to carry out the enzymatic reaction, then it is stopped after an appropriate time and the optical density is measured. of the resulting mixture, if necessary, using a dye or a chromogenic compound (eg o-phenylenediamine). Alternatively, when a fluorescent substance is used to label lectin, a predetermined amount of diluent, such as a buffer solution, is added to the reaction mixture on the gel after incubation, and the intensity of fluorescence is measured. of the resulting mixture.

A procedure which is particularly suitable for implementing the method of the invention uses a kit for the determination of TAGs in a mood of the organism, such as plasma or serum. This kit may include a reagent containing a plant lectin, such as a TAG-specific agglutinating agent. The lectin reagent may also contain a stabilizer and / or a preservative for lectin, such as proteins with a low sugar content, for example beef serum albumin. In a preferred embodiment, this vegetable lectin can be lyophilized and the kit can also include a reconstitution reagent containing an aqueous or water-miscible solvent (pH of approximately 6 to 7.8, preferably 7 7.2). The reagents can optionally also contain buffer substances to maintain the reconstituted reactive system at a set pH and / or stabilizers to prevent deterioration of the substance before use. Although the buffer substances are not considered to be an essential component of the reagents for the kit, a pH of approximately 6 to 7.8, in particular from 7 to 7.2, is preferably used in carrying out the process according to the invention. The reconstitution reagent may preferably contain water as a solvent. For example, a saline solution, a phosphate buffer solution, etc. can be used as a reconstituting agent. In addition, a water-miscible solvent can be added which does not interfere with the reaction.

As indicated above, it is possible advantageously to determine according to the invention the TAG content in a body humor sample. In addition, the invention which makes it possible to recognize the glucosidic bonds associated with tumors can be widely applied to various cancers or tumors, compared to conventional methods or systems using antigenicity and mainly recognizing the protein portion of glucoproteins such as CEA, a1 - fetoprotein, etc., and can be easily applied to the diagnosis of any cancer or tumor, such as stomach cancer, breast cancer, colon cancer, rectal cancer, ovarian cancer, oral cavity cancer, tongue cancer, larynx cancer, prostate cancer, liposarcoma, malignant melanoma, malignant lymphoma, primary sarcoma of the stomach, hepatoma, etc., in their preliminary or advanced stages, and the invention is therefore very useful for the detection of cancers in their initial stages.

The following examples illustrate the invention without, however, limiting its scope.

In these examples, the parts and percentages are all by weight, unless otherwise indicated.

Example 1:

Determination using the blood cell agglutination inhibition reaction i) Preparation of the test sample

Blood is collected from each of seven stomach cancer patients, four lung cancer patients, three breast cancer patients, and patients each with colon cancer, rectal cancer, the ovary, cancer of the oral cavity, cancer of the tongue, cancer of the larynx, cancer of the uterus, cancer of the prostate, malignant melanoma, liposarcoma, malignant lymphoma and primary sarcoma of stomach, as well as in seven healthy people, respectively, and samples are prepared in the conventional manner.

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than. A 6% aqueous solution of sulfosalicylic acid is added in portions to the plasma samples with stirring until the proportion of the plasma additive is 1: 1 by volume. After sufficient agitation, the mixture is centrifuged at 3000-3500 rpm for 10 min and the supernatant is dialyzed with water and 0.15 M phosphate buffer solution for 24 h to prepare the test sample .

ii) Determination

To each of the test samples obtained in i) above, an equal volume of a 0.15 M phosphate buffer solution is added containing in solution 0.8 μg / ml of peanut lectin (produced by the company EY Laboratories Inc.) and the mixture is reacted with stirring at 20-37 ° C for 30 min. The reaction mixture is diluted with a 0.15 M phosphate buffer solution to obtain a series of double dilutions which are each placed on a V plate at the rate of 50 µl and mixed with 50 µl of erythrocytes of rabbit (MO8 at 2-108 cells / ml) with shaking After having allowed the dilution to stand at 37 ° C. for 1 h, the agglutination of the erythrocytes is observed in each dilution. The value of the maximum dilution at which the agglutination still occurs is defined as the agglutination index and the evaluation is carried out using this value The results obtained are shown in the table.

iii) Observations

As is clear from the results shown in the table, all healthy people have an agglutination index of 128 or more, while cancer patients have a much lower agglutination index than healthy patients. Six of the seven patients with stomach cancer have an agglutination index of 8 or less and the rest an index of 32. The four patients with lung cancer have an agglutination index of 1. Regarding other cancers, almost all patients have agglutination indices of 8 or less, except for an index of 32 observed in a patient with uterine cancer and a patient with primary sarcoma stomach.

As indicated above, the agglutination index observed in patients with a malignant tumor or cancer is always 32 or less, while that of healthy people is 128 or more.

The method of the invention is therefore effective in determining the TAG content in a sample of the organism's humor outside the organism and can be useful for the diagnosis of malignant tumors or various cancers.

Example 2:

Determination using the labeled enzyme-lectin complex

Reference stage:

i) Activation of Peroxidase

To 1 ml of an aqueous solution of 0.3M sodium hydrogencarbonate in which 5 mg of peroxidase derived from horseradish was dissolved, 0.1 ml of ethanolic solution of fluorodinitro-benzene 0.1 M is added and the mixture is gently stirred mixing for 1 h at room temperature. 1 ml of 0.06M aqueous NaI04 solution is then added to the mixture which is gently stirred at room temperature for 30 min. The resulting mixture was further stirred with 1 ml of 0.1 M aqueous ethylene glycol solution and stirred gently at room temperature for 1 h, then the reaction mixture was dialyzed with a carbonic acid / hydrogen carbonate buffer solution. 0.1 M sodium (pH 9.5) at 4 ° C for 1 d,

to obtain an activated peroxidase preparation.

ii) Labeling of lectin with Peroxidase

5 mg of lectin (peanut lectin) are dissolved in 3 ml of the peroxidase preparation obtained in i) above and the mixture is gently stirred at room temperature for 2 to 3 h to allow it to react. After adding 5 mg of NaBH4, the reaction mixture is left to stand at 4 ° C for 3 h and the resulting solution is dialyzed with 0.1 M tris-HCl buffer solution (pH 7.4)

for 1 d, then subjected to gel filtration using the

Chromatography on a Sephadex G 150 gel column (eluent: 0.1 M tris-HCl buffer solution at pH 7.4). The optical densities (D28o and D403) of each fraction obtained are measured and the fractions whose peaks of D280 and D403 overlap are collected.

iii) Preparation of agarose gel

Agarose (produced by the company Iwai Kagaku Co., Ltd.) is suspended in a 0.01M tris-HCl buffer solution (pH 7.5), at a rate of 1% by weight, and it is heated the suspension at 70-80 ° C to dissolve it. To this solution is added 0.01% by volume of thimerosal and the resulting solution is poured into test tubes at a rate of 1 ml per tube, then allowed to stand at room temperature to prepare a 1% agarose gel in weight.

Example 2-1:

Polyethylene glycol method i) Preparation of the test sample

5 ml of blood are taken from each of twenty-five cancer patients, two patients with non-cancer diseases and twenty-three healthy people, using a heparin-treated syringe (500 units), and centrifuged at 2000 rpm for 10 min. The resulting supernatant liquid (plasma) is used as the test sample.

ii) Determination

The test sample prepared in i) above is placed in two test tubes at a rate of 200 μl per tube and to each is added 50 ml of the peroxidase-labeled lectin preparation containing 3.5 μg / ml of lectin in a tris-HCl buffer solution (pH about 7.5) prepared in the reference step of Example 2. After having stirred lightly, the mixture is left to stand at 20-30 ° C for 1 h to let him react. 250 μl of a solution of polyethylene glycol (molecular weight 6000) at 8% by weight by volume in a 0.1 M tris-HCl buffer solution are then added to one of the test tubes 250 μl and the mixture is gently stirred to form sample A, and add to the other test tube 250 μl of 0.1M tris-HCl buffer solution and the mixture is gently stirred to form sample B. The two samples A are centrifuged and B for 40 to 60 min using a 1000 G swing type rotor after allowing the sample to stand at 20-30 ° C for 30 to 60 min to allow it to react, collect 50 d of the resulting supernatant and add to 2 ml of physiological saline solution prepared above, after having stirred sufficiently, mix with 500 μl of a substrate solution consisting of 0.1 M orthophenylenediamine citrate buffer solution (final concentration 6% by weight) and 0.1% by weight of hydrogen peroxide and reacted at 20-30 ° C for 30 min in the dark. The substrate solution is prepared beforehand and stored at 4 ° C, which is preferable. Then, 1 ml of 2N hydrochloric acid is added to the reaction mixture to stop the reaction and the color of the sample is measured spectrophotometrically and is expressed by its optical density at 492 nm.

Fig. 1 attached represents the amount of TAG-lectin complex defined as the difference c between the optical density a of sample A and the optical density b of sample B, the symbol 0 representing healthy people and the numbers 1 to 27 denoting the following patients: 1 and 18, stomach cancer; 2, 6, 9, 16, 17, lung cancer; 3, 5, 7, 8 and 13, stomach cancer (preliminary stage); 4 and 14, progressive stomach cancer; 10, heart failure; 11, lymphadenitis; 12, colon cancer with liver metastases; 15, uterine cancer with lung metastases; 19, hydatform mole; 20, colon cancer; 21, uterine cancer; 22 and 26, ovarian cancer; 23, rectal cancer; 24, prostate cancer; 25, cancer of the urethra, and 27, hepatoma.

From the results indicated in fig. 1, it can be seen that the samples having a value c greater than that observed in the samples taken from healthy people contain quantities of TAG greater than those of samples from healthy people. This strongly suggests that people with high c values have tumor or cancer cells.

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Example 2-2:

Gel filtration method i) Blood was collected from fourteen cancer patients and eight healthy people and test samples were prepared in the same manner as in Example 2.

ii) Determination

5 μl of dilution of the test sample which was prepared in i) above are mixed and diluted 104 times with solution H consisting of distilled water containing in solution 8% of sodium chloride, 0.4% of potassium chloride, 0.05% sodium hydrogen phosphate, 0.06% potassium dihydrogen phosphate, 0.05% magnesium sulfate, 0.05% magnesium chloride, 0.1% calcium chloride and 0.1% glucose, with 50 µl of the peroxidase labeled lectin preparation which was prepared in the reference step of Example 2, diluted 100 x with 0.1 M phosphate buffer solution (pH 7.4) and incubated for 15 min in a water bath at 23 ° C. The resulting mixture is incubated for 15 min at 23 ° C, then it is gently applied to the surface of a prepared agarose gel by the method of the reference step of Example 2. After incubation of the test tube containing the gel in a water bath at 23 ° C for 1 h, 2 ml of 0.1M phosphate buffer solution are added (pH about 7 , 4) agarose gel which is then stirred. The supernatant liquid is immediately transferred to a clean test tube, to which 400 μl of substrate solution for the peroxidase prepared in the reference step of Example 2 are then added. After having stirred sufficiently, the mixture is left to stand. mix at room temperature for 15 min to react. The enzymatic reaction is stopped with 1 ml of IN hydrochloric acid and the reaction mixture is sufficiently stirred using a thermomixer or the like. The optical density is measured at 492 nm of the mixture thus treated using a spectrophotometer, a mixture of 400 ml of the substrate solution and 1 ml of IN hydrochloric acid is used as a control. The optical density values obtained are treated in the same way as with example 2-1 and the calculated amount of TAG-lectin is shown in Fig. 2, in which the symbol 0 represents healthy people and the numbers 1 to 10 indicate the following patients: 1, cancer of the es tomac; 2, hydatidiform mole; 3, colon cancer; 4, uterine cancer; 5, ovarian cancer; 6, rectal cancer; 7, prostate cancer; 8, cancer of the urethra; 9, hepatoma, and 10, breast cancer.

As is clear from the results indicated in fig. 2, there is a remarkable difference in the levels of TAG in the body's moods between healthy people and patients with various cancers.

The invention lends itself to practical application in clinical laboratories in the determination of TAG contents in the humors of the organism, such as blood serum or plasma.

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Claims (15)

645,728 1. Method for determining the content of substances containing glucosic linkages associated with tumors (hereinafter designated by the abbreviation TAG) in a mood sample of the organism, characterized in that the TAGs are reacted present in a sample of the organism's mood with a lectin to form a TAG-lectin complex and the amount of TAG-lectin complex or the unreacted lectin is measured. 2. Method according to claim 1, characterized in that a labeled lectin is used. 2 3. Method according to claim 2, characterized in that said labeled lectin is a lectin labeled with an enzyme, a fluorescent substance or a radioactive substance. 4. Method according to claim 2, characterized in that a protective protein is added to said mood of the organism. 5 TAG in a mood sample of the organism and the TAG content obtained is compared with that determined in mood samples of healthy people. 23. Necessary for the implementation of the method according to claim 1, characterized in that it comprises a lectin as agglutinating agent specific for TAG. 24. Kit according to claim 23, characterized in that said lectin has been lyophilized and in that it also contains a reconstitution reagent containing an aqueous or water-miscible solvent. 5. Method according to claim 1, characterized in that said humor of the organism is blood, tissue humor, lymph, hydrothorax, amniotic liquid acid, gastric juice, urine, pancreatic juice, cerebrospinal fluid or saliva. 6. Method according to claim 5, characterized in that said humor of the organism is blood serum or blood plasma. 7. Method according to claim 1, characterized in that the reaction is carried out at 45 ° C or less for at least 30 min. 8. Method according to claim 7, characterized in that the reaction is carried out at 4-40 ° C. 9. Method according to claim 7, characterized in that the reaction is carried out at 20-40 ° C. 10. Method according to claim 7, characterized in that the reaction is carried out for 60 to 90 min. 11. Method according to claim 1, characterized in that said lectin is a lectin which specifically binds on the galac-tose- (ßl -> 3 or ßl -> 4) -N-acetylglucosamine, and / or -acetylgalactos- bond. amine. 12. Method according to claim 11, characterized in that said lectin is peanut lectin or castor seed lectin. 13. The method of claim 1, characterized in that the amount of said unreacted lectin is determined by agglutination of said unreacted lectin. 14. Method according to claim 13, characterized in that the unreacted lectin is agglutinated with a glucoprotein containing a galactose- (ßl - "3 or ßl - <- 4) -N-acetylglucosamine and / or acetylgalactosamine terminal bond. , or with a polymeric derivative of dextran or glass beads coated with said glucoprotein. 15. The method of claim 14, characterized in that said glucoprotein consists of rabbit erythrocytes or sheep erythrocytes treated with neuraminidiase. 16. The method of claim 3, characterized in that said enzyme is glucoamylase, glucose oxidase, peroxidase, alkaline phosphatase or hemeoctapeptide or an active fragment thereof. 17. The method of claim 3, characterized in that said fluorescent substance is fluorescein, fluorescein hydrogenothiocyanate or dansyl chloride. 18. The method of claim 3, characterized in that said radioactive substance is radioactive iodine or tritium. 19. The method of claim 16, characterized in that said enzyme is peroxidase. 20. The method of claim 1, characterized in that one separates a fraction of TAG from the mood of the organism, one reacts this fraction with the marked or unlabeled lectin, one separates the lectin not having reacted from the reaction mixture and the amount of unreacted lectin or lectin reacted with TAGs is measured. 21. The method of claim 1, characterized in that a humor of the organism is reacted with the labeled or unlabeled lectin, the reacted lectin is separated from the unreacted lectin on a gel, and the amount of reactin or unreacted lectin is measured. 22. Application of the method according to claim 1 to the diagnosis of cancer, characterized in that the content of 25. Kit according to claim 23, characterized in that said lectin is a peanut lectin.